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A family of phosphodiesterase inhibitors discovered by cocrystallography and scaffold-based drug design


Cyclic nucleotide phosphodiesterases (PDEs) comprise a large family of enzymes that regulate a variety of cellular processes. We describe a family of potent PDE4 inhibitors discovered using an efficient method for scaffold-based drug design. This method involves an iterative approach starting with low-affinity screening of compounds followed by high-throughput cocrystallography to reveal the molecular basis underlying the activity of the newly identified compounds. Through detailed structural analysis of the interaction of the initially discovered pyrazole carboxylic ester scaffold with PDE4D using X-ray crystallography, we identified three sites of chemical substitution and designed small selective libraries of scaffold derivatives with modifications at these sites. A 4,000-fold increase in the potency of this PDE4 inhibitor was achieved after only two rounds of chemical synthesis and the structural analysis of seven pyrazole derivatives bound to PDE4B or PDE4D, revealing the robustness of this approach for identifying new inhibitors that can be further developed into drug candidates.

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Figure 1: Crystal structure of the pyrazole scaffold and its derivatives in complex with PDE4B or PDE4D.
Figure 2: Correlation of calculated binding energy with experimental IC50 against PDE4B.
Figure 3: Pyrazole scaffold bound to PDE4B and PDE4D and the discovery of potent pyrazole inhibitors for PDE4 in three steps.
Figure 4: Data collection, processing, and refinement statistics for all the cocrystal structures of various pyrazoles in complex with PDE4B or PDE4D.

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We thank Peter Hirth for his insight and inspiration on the scaffold-based drug design paradigm. We thank Axel T. Brunger for critical reading of this manuscript and for discussions. We also thank our colleagues at Plexxikon for help and discussions. We acknowledge Jim Arnold, Fernando Martin and Richard Fronko for the scaffold library design and construction. We thank Ben Powell, Catherine Luu and Hoa Nguyen for protein purification. We thank Jaina Thompson for compound preparation for screening and cocrystallization. We thank Heike Krupka, Abhinav Kumar and Weiru Wang for assisting in crystal mounting and data collection, and Abhinav Kumar for assisting in structure determination. Diffraction data were collected at the Advanced Light Source and the Stanford Synchrotron Radiation Laboratory, which are supported by the US Department of Energy, Office of Basic Energy Sciences under contract DE-AC03-76SF00098 and DE-AC03-76SF00515, respectively. J.S. is supported by National Institutes of Health grant 1RO1-AR051448-01 and funds from the Ludwig Institute for Cancer Research.

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Correspondence to Kam Y J Zhang.

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All the authors except S.-H.K. and J.S. are employees of Plexxikon. S.-H.K. and J.S. are cofounders of the company and shareholders of Plexxikon.

Supplementary information

Supplementary Table 1

Biochemical inhibition data of the pyrazoles against a full panel of PDEs. (PDF 213 kb)

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Card, G., Blasdel, L., England, B. et al. A family of phosphodiesterase inhibitors discovered by cocrystallography and scaffold-based drug design. Nat Biotechnol 23, 201–207 (2005).

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